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Single electron transfer in organic synthesis targeted towards sustainable manufacture
Get PDFSingle electron transfer technology has received considerable interest in organic synthesis in recent years. Single electron transfer gives rise to unconventional modes of reactivity and intermediates, thus serving as a platform for innovative bond constructions, deconstructions and functional group transformations. This is of key importance to the pharmaceutical industry, where the brevity of synthetic routes to small molecules both accelerates drug discovery and drives chemical efficiency in late-stage development. In many cases, single electron transfer chemistry unlocks reaction conditions which are milder, safer and more cost-effective than conventional chemistries. This Thesis investigates two fields of single electron transfer research and contributes new methodology and mechanistic understanding to both. Volume 1 investigates the N-functionalisation of tertiary amines, which is an important method for the elaboration of naturally occuring raw materials into pharmaceutically useful compounds. Two complementary methodologies driven by single electron oxidation were developed. The first method, using visible-light photoredox catalysis, achieved selective benzylic N-CHN₂ functionalisation of N-substituted tetrahydroisoquinolines. The second method, using stable radical cation salts, achieved selective N-CHN₃ functionalisation of trialkylamines. Volume 2 investigates transition metal-free C-H arylation reactions, which are of particular importance to the pharmaceutical industry, given the costly nature of transition metals typically used in catalysis and their long-term sustainability. These reactions are triggered by single electron reduction, effected by a combination of simple alkali metal alkoxide bases and cheap, readily available organic additives. The interplay of the alkoxide and the different organic additives is not fully understood but is key to unlocking milder reaction conditions and broadening substrate scope in these reactions. Comprehensive mechanistic studies were undertaken in this regard.Single electron transfer technology has received considerable interest in organic synthesis in recent years. Single electron transfer gives rise to unconventional modes of reactivity and intermediates, thus serving as a platform for innovative bond constructions, deconstructions and functional group transformations. This is of key importance to the pharmaceutical industry, where the brevity of synthetic routes to small molecules both accelerates drug discovery and drives chemical efficiency in late-stage development. In many cases, single electron transfer chemistry unlocks reaction conditions which are milder, safer and more cost-effective than conventional chemistries. This Thesis investigates two fields of single electron transfer research and contributes new methodology and mechanistic understanding to both. Volume 1 investigates the N-functionalisation of tertiary amines, which is an important method for the elaboration of naturally occuring raw materials into pharmaceutically useful compounds. Two complementary methodologies driven by single electron oxidation were developed. The first method, using visible-light photoredox catalysis, achieved selective benzylic N-CHN₂ functionalisation of N-substituted tetrahydroisoquinolines. The second method, using stable radical cation salts, achieved selective N-CHN₃ functionalisation of trialkylamines. Volume 2 investigates transition metal-free C-H arylation reactions, which are of particular importance to the pharmaceutical industry, given the costly nature of transition metals typically used in catalysis and their long-term sustainability. These reactions are triggered by single electron reduction, effected by a combination of simple alkali metal alkoxide bases and cheap, readily available organic additives. The interplay of the alkoxide and the different organic additives is not fully understood but is key to unlocking milder reaction conditions and broadening substrate scope in these reactions. Comprehensive mechanistic studies were undertaken in this regard
Developing multi degree of freedom control brain computer interface system for spinal cord injury patients
Get PDFBrain computer interface (BCI) is a paradigm that offers an alternative communication channel between neural activity generated in the brain and the user's external environment. BCI decodes the brain activity obtained from an electroencephalogram (EEG) signal and convert this information to a sensible output such as commands to control and communicate with the augmentative and assistive devices. Nevertheless, the majority of the existing BCI system associates with healthy subjects operate based on a combination of multiple limbs and bounded by capability of low dimensional control. Besides that, the acquired results also are not an appropriate platform to infer with the neurologically impaired patients (e.g. spinal cord injury patients). This is probably healthy subjects have full control over their limbs and their EEG signatures show a different pattern. On the other hand, neurologically impaired patients have limited access/control over their limbs and the EEG signatures are affected by the side effects of the prescribed medication, deafferentation and cortical reorganization of brain regions as a function of duration, level and type of disease. This study focuses on the feasibility of developing a multi degree of freedom control BCI system using imagination and intention of movement of a single limb for spinal cord injury (SCI) patients. A pilot study has been conducted on eleven healthy subjects to examine the feasibility of the proposed experimental protocol to record data for implementing the same procedure on SCI patients. In the present study, eighteen SCI patients from Queen Elizabeth National Spinal Injury Unit of the Queen Elizabeth University Hospital voluntarily participated as subjects. The participating subjects have performed and imagined performing right wrist movement towards four centre out directions using a custom made manipulandum triggered by a visual cue whilst EEG, electromyography (EMG) and movement signals are recorded simultaneously through NeuroScanTM Synamp system and CED 1401 (Cambridge Electronic Design). The EEG signal was analysed using signal processing and statistical analysis method. Our findings indicate the detection of Bereitschaft potential 500ms before onset of movement and 500ms after onset of the visual cue. Additionally, there are statistical differences in the relative power withinvithe EEG signal rhythm components namely, delta, theta, alpha beta and gamma bands during imagination and intention of movement towards the four different directions. The significant changes of the estimated relative power of EEG components were extracted as features associated with direction. The features then were normalised, cross validated and dimensionality reduced before being classified using k nearest neighbour (k-NN), fuzzy k nearest neighbour (FKNN) and quadratic discriminant analysis (QDA) classifier. The single trial classification results for motor imagery and motor task by k-NN, FKNN and QDA classifier dwell within the range of 52.31%-94.14% and 52.20%-96.51%, respectively. These findings proved that it is possible to develop a functional multi degree of freedom BCI system that employs imagination/intention of movement using a single limb for the SCI population. On top of that the developed BCI system and classification also required no subject training at all.Brain computer interface (BCI) is a paradigm that offers an alternative communication channel between neural activity generated in the brain and the user's external environment. BCI decodes the brain activity obtained from an electroencephalogram (EEG) signal and convert this information to a sensible output such as commands to control and communicate with the augmentative and assistive devices. Nevertheless, the majority of the existing BCI system associates with healthy subjects operate based on a combination of multiple limbs and bounded by capability of low dimensional control. Besides that, the acquired results also are not an appropriate platform to infer with the neurologically impaired patients (e.g. spinal cord injury patients). This is probably healthy subjects have full control over their limbs and their EEG signatures show a different pattern. On the other hand, neurologically impaired patients have limited access/control over their limbs and the EEG signatures are affected by the side effects of the prescribed medication, deafferentation and cortical reorganization of brain regions as a function of duration, level and type of disease. This study focuses on the feasibility of developing a multi degree of freedom control BCI system using imagination and intention of movement of a single limb for spinal cord injury (SCI) patients. A pilot study has been conducted on eleven healthy subjects to examine the feasibility of the proposed experimental protocol to record data for implementing the same procedure on SCI patients. In the present study, eighteen SCI patients from Queen Elizabeth National Spinal Injury Unit of the Queen Elizabeth University Hospital voluntarily participated as subjects. The participating subjects have performed and imagined performing right wrist movement towards four centre out directions using a custom made manipulandum triggered by a visual cue whilst EEG, electromyography (EMG) and movement signals are recorded simultaneously through NeuroScanTM Synamp system and CED 1401 (Cambridge Electronic Design). The EEG signal was analysed using signal processing and statistical analysis method. Our findings indicate the detection of Bereitschaft potential 500ms before onset of movement and 500ms after onset of the visual cue. Additionally, there are statistical differences in the relative power withinvithe EEG signal rhythm components namely, delta, theta, alpha beta and gamma bands during imagination and intention of movement towards the four different directions. The significant changes of the estimated relative power of EEG components were extracted as features associated with direction. The features then were normalised, cross validated and dimensionality reduced before being classified using k nearest neighbour (k-NN), fuzzy k nearest neighbour (FKNN) and quadratic discriminant analysis (QDA) classifier. The single trial classification results for motor imagery and motor task by k-NN, FKNN and QDA classifier dwell within the range of 52.31%-94.14% and 52.20%-96.51%, respectively. These findings proved that it is possible to develop a functional multi degree of freedom BCI system that employs imagination/intention of movement using a single limb for the SCI population. On top of that the developed BCI system and classification also required no subject training at all
Low-complexity low-rate residential non-intrusive appliance load monitoring
Get PDFLarge-scale smart metering deployments and energy saving targets across the world have ignited renewed interest in residential non-intrusive appliance load monitoring (NALM), that is, disaggregating total household's energy consumption down to individual appliances, using purely analytical tools.;Despite increased research efforts, NALM techniques that can disaggregate power loads at low sampling rates are still not accurate and/or practical enough, requiring substantial customer input and long training periods. In this thesis, we address these challenges via a practical low complexitylow-rate NALM, by proposing two approaches based on a combination of the following machine learning techniques: k-means clustering and Support Vector Machine, exploiting their strengths and addressing their individual weaknesses.;The first proposed supervised approach is a low-complexity method that requires very short training period and is robust to labelling errors. The second, unsupervised approach relies on a database of appliance signatures that we designed using publicly available datasets.;The database compactly represents over 100 appliances using statistical modelling of measured active power. Experimental results on three datasets from US (REDD), Italy and Austria (GREEND) and UK (REFIT), demonstrate the reliability and practicality of the proposed approaches.Large-scale smart metering deployments and energy saving targets across the world have ignited renewed interest in residential non-intrusive appliance load monitoring (NALM), that is, disaggregating total household's energy consumption down to individual appliances, using purely analytical tools.;Despite increased research efforts, NALM techniques that can disaggregate power loads at low sampling rates are still not accurate and/or practical enough, requiring substantial customer input and long training periods. In this thesis, we address these challenges via a practical low complexitylow-rate NALM, by proposing two approaches based on a combination of the following machine learning techniques: k-means clustering and Support Vector Machine, exploiting their strengths and addressing their individual weaknesses.;The first proposed supervised approach is a low-complexity method that requires very short training period and is robust to labelling errors. The second, unsupervised approach relies on a database of appliance signatures that we designed using publicly available datasets.;The database compactly represents over 100 appliances using statistical modelling of measured active power. Experimental results on three datasets from US (REDD), Italy and Austria (GREEND) and UK (REFIT), demonstrate the reliability and practicality of the proposed approaches
Vortex-induced vibration of cylindrical structures
Get PDFVortex-induced vibration (VIV) of cylindrical structures is a classical topic within fluid-structure interaction (FSI). In offshore engineering, it often causes the fatigue of slender structures, such as risers, mooring lines and pipelines. Detailed understanding of this FSI phenomenon and an efficient prediction of such self-excited and self-sustained oscillations are required for the reliable estimation of the fatigue damage and the development of VIV suppression techniques.Over the past few decades, VIV has been extensively studied and the majority of the existing publications in the literature are experiments or semi-empirical modelling. In contrast, FSI simulations by combining high-fidelity computational fluid dynamics (CFD) and computational structural dynamics (CSD) solvers have received less attention. The main objective of this thesis is to investigate VIV of elastically mounted rigid cylinders and flexible cylinders using fully three-dimensional (3D) FSI simulations. Apart from important VIV aspects, such as response amplitude, response frequency and fatigue damage etc., the present research is also focussed on the aspects which have not been fully addressed by previous studies such as correlation lengths and time-dependent 3D flow structures.Two-degree-of-freedom (2DOF) VIV of an elastically mounted circular cylinder with varying in-line (IL) to cross-flow (CF) natural frequency ratios (f* = fnx/fny) is first studied using a 3D CFD approach. Numerical simulation is carried out for a constant mass ratio m* = 2 at a fixed Reynolds number Re = 500. The reduced velocity Vr ranges from 2 to 12. Three natural frequency ratios are considered, i.e., f* = 1, 1.5 and 2. The structural damping is set to zero to maximise the response of the cylinder. The main objective of the first study is to investigate the effect of f* on the 2DOF VIV responses and the 3D characteristics of the flow. It is discovered that there is a significant increase in the vibration amplitude and the peak amplitude shifts to a higher reduced velocity when f* increases from 1 to 2. A single-peak cross-flow response is observed for the identical in-line and cross-flow mass ratios when f* = 2. Dual resonance is found to exist over the range of f* studied.;The preferable trajectories of the cylinder in the lock-in range are counterclockwise figure-eight orbits, whereas clockwise orbits primarily occur in the initial branch. The number of clockwise orbits decreases as f* increases from 1 to 2. Oblique figure-eight trajectories appear at Vr = 6, 7 and 8 when f* = 1. The third harmonic component which is observed in the lift fluctuation increases with f*. The correlation decreases in the lock-in range and reaches its minimum value around the transition region between the lock-in and post-lock-in ranges. Three vortex shedding modes (2S, P + S and 2P) appear in the present simulation. A dominant P + S mode is associated with the oblique figure-eight trajectories. Variation of vortex shedding flows along the cylinder is observed leading to the poor correlation of the sectional lift forces.Then, a numerical investigation of VIV of a vertical riser subject to uniform and linearly sheared currents is presented. The model vertical riser tested at the MARINTEK by ExxonMobil is considered. The predicted numerical results are in good agreement with the experimental data. It is found that the dominant mode numbers, the maximum root mean square amplitudes, the dominant frequencies and the fatigue damage indices increase with the flow velocity. Dual resonance is found to occur at most of the locations along the riser. At some locations along the riser, a third harmonic frequency component is observed in the CF response and a frequency component at the CF response frequency is found in the IL response apart from the frequency component at twice the CF response frequency. The majority of the vortex shedding shows a clear 2S pattern, whereas a 2P mode is observed near the position where the maximum vibration amplitude appears. The higher IL fatigue damage in the second study emphasises the importance of the IL fatigue damage analysis especially in the design of low flow velocity or low mode number applications.The third study is on VIV of two tandem flexible cylinders at different spacing ratios (Sx/D) at a fixed Reynolds number Re = 500 using a two-way FSI method. The main objective is to investigate the effect of spacing on the hydrodynamic interactions and the VIV responses of these cylinders. It is found that the responses of the two tandem flexible cylinders are similar to the classical VIV responses when Sx/D is small.;Once Sx/D is large enough for the vortices to be completely detached from the upstream cylinder, the response of the upstream cylinder is similar to the typical VIV response whereas the downstream cylinder undergoes wake-induced vibration (WIV). The characteristics of the response of the downstream cylinder in the present study are similar to those of the first two response regimes. The third response regime is not observed for the flexible downstream cylinder with both ends fixed. The two changes in the phase relation between the cross-flow displacements of the two tandem flexible cylinders are discovered to be linked with the initial-upper branch transition and the upper-lower branch transition, respectively. The correlation lengths of the two tandem flexible cylinders decrease significantly in the transition range between the upper and lower branches. Three vortex shedding modes (2S, P + S and 2P) have been identified in the present study. It is found that the upper-branch 2P mode is associated with large-amplitude vibration of the upstream cylinder and the P + S mode is related to large-amplitude vibration of the downstream cylinder for Sx/D = 3.5 and 5. On the other hand, the lower-branch 2P mode leads to small-amplitude vibration of the downstream cylinder in the post-lock-in range at Sx/D = 2.5. The relative phase shifts of the sectional lift coefficients on different spanwise cross sections can be attributed to the variation of the vortex shedding flow along the flexible cylinders and these phase shifts result in poor phasing between the forces and the displacements which is related to the decrease of the correlation lengths.Vortex-induced vibration (VIV) of cylindrical structures is a classical topic within fluid-structure interaction (FSI). In offshore engineering, it often causes the fatigue of slender structures, such as risers, mooring lines and pipelines. Detailed understanding of this FSI phenomenon and an efficient prediction of such self-excited and self-sustained oscillations are required for the reliable estimation of the fatigue damage and the development of VIV suppression techniques.Over the past few decades, VIV has been extensively studied and the majority of the existing publications in the literature are experiments or semi-empirical modelling. In contrast, FSI simulations by combining high-fidelity computational fluid dynamics (CFD) and computational structural dynamics (CSD) solvers have received less attention. The main objective of this thesis is to investigate VIV of elastically mounted rigid cylinders and flexible cylinders using fully three-dimensional (3D) FSI simulations. Apart from important VIV aspects, such as response amplitude, response frequency and fatigue damage etc., the present research is also focussed on the aspects which have not been fully addressed by previous studies such as correlation lengths and time-dependent 3D flow structures.Two-degree-of-freedom (2DOF) VIV of an elastically mounted circular cylinder with varying in-line (IL) to cross-flow (CF) natural frequency ratios (f* = fnx/fny) is first studied using a 3D CFD approach. Numerical simulation is carried out for a constant mass ratio m* = 2 at a fixed Reynolds number Re = 500. The reduced velocity Vr ranges from 2 to 12. Three natural frequency ratios are considered, i.e., f* = 1, 1.5 and 2. The structural damping is set to zero to maximise the response of the cylinder. The main objective of the first study is to investigate the effect of f* on the 2DOF VIV responses and the 3D characteristics of the flow. It is discovered that there is a significant increase in the vibration amplitude and the peak amplitude shifts to a higher reduced velocity when f* increases from 1 to 2. A single-peak cross-flow response is observed for the identical in-line and cross-flow mass ratios when f* = 2. Dual resonance is found to exist over the range of f* studied.;The preferable trajectories of the cylinder in the lock-in range are counterclockwise figure-eight orbits, whereas clockwise orbits primarily occur in the initial branch. The number of clockwise orbits decreases as f* increases from 1 to 2. Oblique figure-eight trajectories appear at Vr = 6, 7 and 8 when f* = 1. The third harmonic component which is observed in the lift fluctuation increases with f*. The correlation decreases in the lock-in range and reaches its minimum value around the transition region between the lock-in and post-lock-in ranges. Three vortex shedding modes (2S, P + S and 2P) appear in the present simulation. A dominant P + S mode is associated with the oblique figure-eight trajectories. Variation of vortex shedding flows along the cylinder is observed leading to the poor correlation of the sectional lift forces.Then, a numerical investigation of VIV of a vertical riser subject to uniform and linearly sheared currents is presented. The model vertical riser tested at the MARINTEK by ExxonMobil is considered. The predicted numerical results are in good agreement with the experimental data. It is found that the dominant mode numbers, the maximum root mean square amplitudes, the dominant frequencies and the fatigue damage indices increase with the flow velocity. Dual resonance is found to occur at most of the locations along the riser. At some locations along the riser, a third harmonic frequency component is observed in the CF response and a frequency component at the CF response frequency is found in the IL response apart from the frequency component at twice the CF response frequency. The majority of the vortex shedding shows a clear 2S pattern, whereas a 2P mode is observed near the position where the maximum vibration amplitude appears. The higher IL fatigue damage in the second study emphasises the importance of the IL fatigue damage analysis especially in the design of low flow velocity or low mode number applications.The third study is on VIV of two tandem flexible cylinders at different spacing ratios (Sx/D) at a fixed Reynolds number Re = 500 using a two-way FSI method. The main objective is to investigate the effect of spacing on the hydrodynamic interactions and the VIV responses of these cylinders. It is found that the responses of the two tandem flexible cylinders are similar to the classical VIV responses when Sx/D is small.;Once Sx/D is large enough for the vortices to be completely detached from the upstream cylinder, the response of the upstream cylinder is similar to the typical VIV response whereas the downstream cylinder undergoes wake-induced vibration (WIV). The characteristics of the response of the downstream cylinder in the present study are similar to those of the first two response regimes. The third response regime is not observed for the flexible downstream cylinder with both ends fixed. The two changes in the phase relation between the cross-flow displacements of the two tandem flexible cylinders are discovered to be linked with the initial-upper branch transition and the upper-lower branch transition, respectively. The correlation lengths of the two tandem flexible cylinders decrease significantly in the transition range between the upper and lower branches. Three vortex shedding modes (2S, P + S and 2P) have been identified in the present study. It is found that the upper-branch 2P mode is associated with large-amplitude vibration of the upstream cylinder and the P + S mode is related to large-amplitude vibration of the downstream cylinder for Sx/D = 3.5 and 5. On the other hand, the lower-branch 2P mode leads to small-amplitude vibration of the downstream cylinder in the post-lock-in range at Sx/D = 2.5. The relative phase shifts of the sectional lift coefficients on different spanwise cross sections can be attributed to the variation of the vortex shedding flow along the flexible cylinders and these phase shifts result in poor phasing between the forces and the displacements which is related to the decrease of the correlation lengths
Facial analytics for emotional state recognition
Get PDFFor more than 75 years, social scientists study the human emotions. Whereas numerous theories developed about the provenance and number of basic emotions, most agreed that they could categorize into six categories: angrer, disgust, fear, joy, sadness and surprise. To evaluate emotions, psychologists focused their research in facial expressions analysis. In recent years, the progress in digital technologies field has steered the researchers in psychology, computer science, linguistics, neuroscience, and related disciplines towards the usage of computer systems that analyze and detect the human emotions. Usually, these algorithms are referred in the literature as facial emotion recognition (FER) systems. In this thesis, two different approaches are described and evaluated in order to recognize the six basic emotions automatically from still images.An effective face detection scheme, based on color techniques and the well-known Viola and Jones (VJ) algorithm is proposed for the face and facial characteristics localization within an image. A novel algorithm which exploits the eyes' centers coordinates, is applied on the image to align the detected face. In order to reduce the effects of illumination, homomorphic filtering is applied on the face area. Three regions (mouth, eyes and glabella) are localized and further processed for texture analysis.Although many methods have been proposed in the literature to recognize the emotion from the human face, they are not designed to be able to handle partial occlusions and multiple faces. Therefore, a novel algorithm that extracts information through texture analysis, from each region of interest, is evaluated. Two popular techniques (histograms of oriented gradients and local binary patterns) are utilized to perform texture analysis in the abovementioned facial patches. By evaluating several combinations of their principal parameters and two classification techniques (support vector machine and linear discriminant analysis), three classifiers are proposed. These three models are enabled depending on the regions' availability. Although both classification approaches have shown impressive results, LDA proved to be slightly better especially regarding the amount of data management. Therefore, the final models, which utilized for comparison purpose, were trained using LDA classification.Experiments using Cohn-Kanade plus (CK+) and Amsterdam Dynamic Facial Expression Set (ADFES) datasets demonstrate that the presented FER algorithm has surpassed other significant FER systems in terms of processing time and accuracy. The evaluation of the system involved three experiments: intra-testing experiment (train and test with the same dataset), train/test process between CK+ and ADFES and finally the development of a new database based on selfie-photos, which is tested on the pre-trained models. The last two experiments constitute a certain evidence that Emotion Recognition System (ERS) can operate under various pose and light circumstances.For more than 75 years, social scientists study the human emotions. Whereas numerous theories developed about the provenance and number of basic emotions, most agreed that they could categorize into six categories: angrer, disgust, fear, joy, sadness and surprise. To evaluate emotions, psychologists focused their research in facial expressions analysis. In recent years, the progress in digital technologies field has steered the researchers in psychology, computer science, linguistics, neuroscience, and related disciplines towards the usage of computer systems that analyze and detect the human emotions. Usually, these algorithms are referred in the literature as facial emotion recognition (FER) systems. In this thesis, two different approaches are described and evaluated in order to recognize the six basic emotions automatically from still images.An effective face detection scheme, based on color techniques and the well-known Viola and Jones (VJ) algorithm is proposed for the face and facial characteristics localization within an image. A novel algorithm which exploits the eyes' centers coordinates, is applied on the image to align the detected face. In order to reduce the effects of illumination, homomorphic filtering is applied on the face area. Three regions (mouth, eyes and glabella) are localized and further processed for texture analysis.Although many methods have been proposed in the literature to recognize the emotion from the human face, they are not designed to be able to handle partial occlusions and multiple faces. Therefore, a novel algorithm that extracts information through texture analysis, from each region of interest, is evaluated. Two popular techniques (histograms of oriented gradients and local binary patterns) are utilized to perform texture analysis in the abovementioned facial patches. By evaluating several combinations of their principal parameters and two classification techniques (support vector machine and linear discriminant analysis), three classifiers are proposed. These three models are enabled depending on the regions' availability. Although both classification approaches have shown impressive results, LDA proved to be slightly better especially regarding the amount of data management. Therefore, the final models, which utilized for comparison purpose, were trained using LDA classification.Experiments using Cohn-Kanade plus (CK+) and Amsterdam Dynamic Facial Expression Set (ADFES) datasets demonstrate that the presented FER algorithm has surpassed other significant FER systems in terms of processing time and accuracy. The evaluation of the system involved three experiments: intra-testing experiment (train and test with the same dataset), train/test process between CK+ and ADFES and finally the development of a new database based on selfie-photos, which is tested on the pre-trained models. The last two experiments constitute a certain evidence that Emotion Recognition System (ERS) can operate under various pose and light circumstances
Developing higher-order reading skills in mainstream primary schools : a metacognitive approach
Get PDFResearch indicates that multiple strategy comprehension instruction (MSCI) programmes in general yield greater effect sizes than single strategy approaches. Potential candidate MSCI interventions were evaluated on the basis of effect size of outcome, feasibility and acceptability, and universality. This identified the Strathclyde Higher-Order Reading Skills (SHORS) Programme as a promising intervention yet to be evaluated in the UK in a controlled study. Following a pilot study, a mixed-model quasi-experimental study was carried out with condition (intervention versus control group) and reading comprehension ability (higher versus average versus lower tertiles) as between-group independent variables and time-point (pre- versus post-intervention) as a within-subject independent variable. The WIAT-IIUK reading comprehension subtest was the primary outcome measure. Seventy-four pupils in five Primary 5 classes (aged 9-10) in four primary schools were recruited as participants from within a Scottish local authority. Training and implementation of the SHORS intervention followed the procedure of McCartney, Boyle & Ellis (2015) study, with delivery of 4 sessions of 45 minutes per week for 8 weeks. Comparison of pre and post reading comprehension scores showed a statistically significant intervention effect (Cohen's d = 0.81), which exceeded the minimally significant difference of d = 0.67 taking precision of measurement and measurement error into account (Weir, 2005). Participants in the higher, average and lower tertiles of pre-intervention reading comprehension scores all benefited equally, indicating that the SHORS may be regarded as a 'universal' intervention. Secondary quantitative and qualitative data confirm that the intervention is easy to implement, feasible within a Scottish setting and acceptable to school staff.The study extends the reading intervention literature regarding learner, teacher and learning environment, implementation, metacognitive knowledge and reading habits. Implications for policy and practice are discussed.Research indicates that multiple strategy comprehension instruction (MSCI) programmes in general yield greater effect sizes than single strategy approaches. Potential candidate MSCI interventions were evaluated on the basis of effect size of outcome, feasibility and acceptability, and universality. This identified the Strathclyde Higher-Order Reading Skills (SHORS) Programme as a promising intervention yet to be evaluated in the UK in a controlled study. Following a pilot study, a mixed-model quasi-experimental study was carried out with condition (intervention versus control group) and reading comprehension ability (higher versus average versus lower tertiles) as between-group independent variables and time-point (pre- versus post-intervention) as a within-subject independent variable. The WIAT-IIUK reading comprehension subtest was the primary outcome measure. Seventy-four pupils in five Primary 5 classes (aged 9-10) in four primary schools were recruited as participants from within a Scottish local authority. Training and implementation of the SHORS intervention followed the procedure of McCartney, Boyle & Ellis (2015) study, with delivery of 4 sessions of 45 minutes per week for 8 weeks. Comparison of pre and post reading comprehension scores showed a statistically significant intervention effect (Cohen's d = 0.81), which exceeded the minimally significant difference of d = 0.67 taking precision of measurement and measurement error into account (Weir, 2005). Participants in the higher, average and lower tertiles of pre-intervention reading comprehension scores all benefited equally, indicating that the SHORS may be regarded as a 'universal' intervention. Secondary quantitative and qualitative data confirm that the intervention is easy to implement, feasible within a Scottish setting and acceptable to school staff.The study extends the reading intervention literature regarding learner, teacher and learning environment, implementation, metacognitive knowledge and reading habits. Implications for policy and practice are discussed
Mechanistic pharmacokinetic / pharmacodynamic modelling of acute inflammatory challenge models
Get PDFPreviously held under moratorium in Chemistry Department (GSK) from 31st May 2017 to 31st May 2019.The confidentiality statement on each page of this thesis DOES NOT applySystemic Lupus Erythematosus (SLE), is a chronic autoimmune disease that can affectmultiple organ systems. Survival rates have improved in recent years but SLE is stillcurrently “managed” rather than “cured” and traditional treatments can cause irreversibledamage, sometimes leading to death. Consequently, there is a need for new therapeuticagents that target specific components of the disease pathogenesis. This has provedchallenging since the exact aetiology of SLE is undefined, although most of the cells of theimmune system have been implicated in the disease process.The cytokine interferon (IFN) α has a key role in the early induction of the disease andongoing pathogenesis. Immune complexes of autoantigens and autoantibodies have beenfound in the sera of SLE patients and induce the production of IFNα by plasmacytoiddendritic cells (pDCs) through toll like receptors. IFNα binds to a specific receptor whichresults in the regulation of genes associated with the viral response (the IFN genesignature). This signature has been correlated with disease activity in SLE patientsreinforcing a role of IFNα in SLE. IFNα can also stimulate and inhibit a wide range ofcytokines. In SLE, a vicious circle is established with an ongoing production of IFNα frompDCs that maintains the autoimmune process.Lupus mouse models that mimic a human SLE like disease have previously been used toinvestigate new treatments for SLE, but have only delivered drugs that treat the symptomsrather than modifying the disease. An alternative approach is to use mechanism basedmodels to investigate the role of a particular pathway or disease mechanism. PK/PDmodelling of the data arising from challenge models can provide confidence that theefficacy observed in preclinical studies can be translated to the clinical setting and may improve the overall efficiency and success of drug discovery programmes. The IFNαpathway looks a promising target for new treatments for SLE and the induction of IFNα and related biomarkers appear to be translatable between preclinical and clinical species. Therefore this looks an appropriate pathway to investigate using mechanistic preclinica lPK/PD challenge models where the selected challenge agent would induces elements of the IFNα pathway under investigation. Aims The aim of this project was to take a disease focused approach to select and investigate two preclinical mechanistic acute in vivo PK/PD models. These models were selected based onthe hypothesis that the IFNα pathway has a key role in the pathogenesis of SLE. Methods This project investigated two preclinical mechanistic in vivo PK/PD challenge models with challenge agents that stimulate different components of IFNα pathway. One was a highthroughput rodent model designed to enable screening of a number of compounds whilethe other was a primate model that was considered translatable to human. Mouse model A mouse model was investigated that used the small molecule TLR7 agonist resiquimod to induce IFNα production from pDCs. First, the pharmacokinetics of resiquimod were determined after iv infusion of 0.25 mg/kg. The relationship between IFNα response and dose was then determined over the range 0.04 to 4.8 mg/kg. Finally, a fixed dose of 0.4mg/kg was used to determine the reproducibility of the IFNα response between study days. Concentrations of resiquimod and IFNα were determined by LC-MSMS and ELISA analysis respectively. The data were initially analysed by non-compartmental analysis to determine the pharmacokinetics of resiquimod. Statistical and power analysis were then applied to determine the reproducibility of the model between study days and to understand the potential utility of the model in the drug discovery. Finally, a population PK/PD analysis of the dose response relationship was conducted using a range of models that included anendogenous modulator function to describe the inhibition of excessive IFNα production. Primate model A primate model was investigated that used recombinant human IFNα2b (INTRON A) to investigate the induction of a range of biomarkers downstream of the IFN receptor. Cynonologus monkeys received a subcutaneous administration of vehicle and IFNα2b at both 3 MIU/kg and 10 MIU/kg. The pharmacokinetics of IFNα2b were determined and the induction of 29 cytokines/chemokines, neopterin and body temperature was investigated. Concentrations of IFNα2b and other cytokines/chemokines were determined by Milliplex® magnetic bead panel, concentrations of neopterin were determined by ELISA analysis and body temperature was determined using a rectal thermometer. The data were initially analysed by non-compartmental analysis to determine the pharmacokinetics of IFNα2b. A population PK/PD analysis of the IFNα2b induction of 6biomarkers was then conducted using an indirect response model with stimulation of input. Results Mouse model Following iv infusion, resiquimod had a blood clearance of 69 mL/min/kg, a volume of distribution at steady state of 2.4 L/kg and a terminal half-life of 0.5 h. There was low interanimal variability in pharmacokinetic parameters and similar results were obtained for iv infusion and iv bolus administration. Linearity in pharmacokinetics was demonstrated over a 120 fold dose range. Following iv administration of resiquimod, IFNα concentrations were observed from 0.75 hpost dose, Cmax occurred at 1-2 h and the last measurable concentration was observed at approximately 3 h. A bell shaped dose response curve was observed with the maximum response observed at 0.09 mg/kg. A dose of 0.4 mg/kg achieved reproducible response results across four study days and was recommended as the challenge dose for future studies. Power analysis demonstrated that the model could be used to investigate multiple compounds at a single dose and the dose response of a single compound. Population PK/PD modelling was conducted in a sequential manner. The pharmacokinetics of resiquimod were described with a 1 compartment iv bolus model with IIV on clearance. The final PK/PD model comprised an indirect response model with stimulation of input, an effect compartment and endogenous modulator function with fixed parameter estimates for Keo, the Hill co-efficient, and M50 and IIV on Keo, EC50 and Emax. The final model did not adequately predict the IFNα time profile in individual mice and highlights that increased data may be required to provide robust estimates of the induction and elimination phase of the IFNα response. The final model did characterise the dose response relationship but predicted a reduction in response at higher doses of resiquimod when IIV was included on Emax rather than the modulator function. Primate model Following sc administration, IFNα2b had an apparent blood clearance of 2.9 mL/min/kg, an apparent volume of distribution of 3.1 L/kg and a terminal half-life of 13 h. There was low inter-animal variability in the systemic exposure (CV of 25-40%) and clearance (CV of 22-37%), however the volume and half-life demonstrated greater variability (CV of >100%)Linearity in pharmacokinetics was demonstrated over a 3 fold dose range. The pharmacokinetics were broadly comparable to those previously reported in the literature. Following IFNα2b treatment induction of neopterin and the cytokines/chemokines IL15,IL1Ra, MCP1, IL6 and eotaxin was observed. There was a sub proportional increase in the concentrations of biomarkers with the increase of IFNα2b dose from 3 MIU/kg to 10MIU/kg. An induction of body temperature was not observed. Population PK/PD modelling was conducted in a sequential manner. The pharmacokineticsof IFNα2b were described with a 2 compartment model with first order absorption and IIVon clearance and IOV on volume. A covariate analysis indicated there was no relationship with either clearance or volume with body weight. The final PK/PD model for all 6biomarkers comprised an indirect response model with stimulation of input and IIV on baseline. In addition IIV on Emax was included for neopterin, IL15, IL6 and MCP1, on Kout forIL15 and EC50 for IL1Ra and eotaxin. High values for IIV were determined for IL6 and IL1Raand the parameters for MCP1 and IL6 demonstrated poor precision. The model did not give a robust prediction of EC50 for any biomarker or Emax for IL6, MCP1 and eotaxin due to the investigated doses giving a comparable response. Conclusions Mouse Model The pharmacokinetics of resiquimod were successfully determined in the mouse for the first time following both iv infusion and iv bolus administration of resiquimod. The investigation of the dose response relationship delivered comparable data to that previously reported in the literature whereby the greatest response was observed at low doses of resiquimod and further increase in dose results in an apparent reduction in the IFNα response to a plateau. A reproducible IFNα response can be achieved between mice receiving the same resiquimod treatment across multiple study days and suggests that model that may be used in drug discovery to investigate therapeutics that have an action on the IFNα pathway. Population PK/PD modelling with the incorporation of anendogenous modulator has highlighted the gaps in the data set and improved the understanding of the model that can be used to guide future PK/PD modelling efforts. Monkey Model The pharmacokinetics of IFNα2b demonstrated that at the doses investigated in this study the receptor mediated clearance had become saturated and the clearance was driven by renal elimination and catabolism. The induction of serum neopterin concentrations andcytokines/chemokines IL15, IL6, MCP1, eotaxin and IL1Ra, which have all been implicated inthe pathogenesis of SLE, was observed in the challenged primates. A less than proportional increase in biomarker concentrations was observed with the increase in dose indicating that the concentrations of IFNα2b associated with these doses may be near the top of the concentration response curve. A population PK/PD model that describes the induction of the 6 biomarkers was developed. However, the model was limited by the small number of doses investigated. Due to the translatability of the biology between primate and humans this project has delivered a PK/PD model with disease relevant endpoints that can be used to screen compounds and potentially predict efficacious clinical doses. This project has successfully designed and validated two preclinical in vivo mechanistic PK/PD challenge models based on the hypothesis that the cytokine IFNα is central to the pathogenesis in SLE and that the mechanisms behind the induced response are translatable between species. This project has demonstrated that both models have potential in drug discovery to be used as tools to select the most appropriate compounds for progression to the clinic and to predict efficacious doses.Systemic Lupus Erythematosus (SLE), is a chronic autoimmune disease that can affectmultiple organ systems. Survival rates have improved in recent years but SLE is stillcurrently “managed” rather than “cured” and traditional treatments can cause irreversibledamage, sometimes leading to death. Consequently, there is a need for new therapeuticagents that target specific components of the disease pathogenesis. This has provedchallenging since the exact aetiology of SLE is undefined, although most of the cells of theimmune system have been implicated in the disease process.The cytokine interferon (IFN) α has a key role in the early induction of the disease andongoing pathogenesis. Immune complexes of autoantigens and autoantibodies have beenfound in the sera of SLE patients and induce the production of IFNα by plasmacytoiddendritic cells (pDCs) through toll like receptors. IFNα binds to a specific receptor whichresults in the regulation of genes associated with the viral response (the IFN genesignature). This signature has been correlated with disease activity in SLE patientsreinforcing a role of IFNα in SLE. IFNα can also stimulate and inhibit a wide range ofcytokines. In SLE, a vicious circle is established with an ongoing production of IFNα frompDCs that maintains the autoimmune process.Lupus mouse models that mimic a human SLE like disease have previously been used toinvestigate new treatments for SLE, but have only delivered drugs that treat the symptomsrather than modifying the disease. An alternative approach is to use mechanism basedmodels to investigate the role of a particular pathway or disease mechanism. PK/PDmodelling of the data arising from challenge models can provide confidence that theefficacy observed in preclinical studies can be translated to the clinical setting and may improve the overall efficiency and success of drug discovery programmes. The IFNαpathway looks a promising target for new treatments for SLE and the induction of IFNα and related biomarkers appear to be translatable between preclinical and clinical species. Therefore this looks an appropriate pathway to investigate using mechanistic preclinica lPK/PD challenge models where the selected challenge agent would induces elements of the IFNα pathway under investigation. Aims The aim of this project was to take a disease focused approach to select and investigate two preclinical mechanistic acute in vivo PK/PD models. These models were selected based onthe hypothesis that the IFNα pathway has a key role in the pathogenesis of SLE. Methods This project investigated two preclinical mechanistic in vivo PK/PD challenge models with challenge agents that stimulate different components of IFNα pathway. One was a highthroughput rodent model designed to enable screening of a number of compounds whilethe other was a primate model that was considered translatable to human. Mouse model A mouse model was investigated that used the small molecule TLR7 agonist resiquimod to induce IFNα production from pDCs. First, the pharmacokinetics of resiquimod were determined after iv infusion of 0.25 mg/kg. The relationship between IFNα response and dose was then determined over the range 0.04 to 4.8 mg/kg. Finally, a fixed dose of 0.4mg/kg was used to determine the reproducibility of the IFNα response between study days. Concentrations of resiquimod and IFNα were determined by LC-MSMS and ELISA analysis respectively. The data were initially analysed by non-compartmental analysis to determine the pharmacokinetics of resiquimod. Statistical and power analysis were then applied to determine the reproducibility of the model between study days and to understand the potential utility of the model in the drug discovery. Finally, a population PK/PD analysis of the dose response relationship was conducted using a range of models that included anendogenous modulator function to describe the inhibition of excessive IFNα production. Primate model A primate model was investigated that used recombinant human IFNα2b (INTRON A) to investigate the induction of a range of biomarkers downstream of the IFN receptor. Cynonologus monkeys received a subcutaneous administration of vehicle and IFNα2b at both 3 MIU/kg and 10 MIU/kg. The pharmacokinetics of IFNα2b were determined and the induction of 29 cytokines/chemokines, neopterin and body temperature was investigated. Concentrations of IFNα2b and other cytokines/chemokines were determined by Milliplex® magnetic bead panel, concentrations of neopterin were determined by ELISA analysis and body temperature was determined using a rectal thermometer. The data were initially analysed by non-compartmental analysis to determine the pharmacokinetics of IFNα2b. A population PK/PD analysis of the IFNα2b induction of 6biomarkers was then conducted using an indirect response model with stimulation of input. Results Mouse model Following iv infusion, resiquimod had a blood clearance of 69 mL/min/kg, a volume of distribution at steady state of 2.4 L/kg and a terminal half-life of 0.5 h. There was low interanimal variability in pharmacokinetic parameters and similar results were obtained for iv infusion and iv bolus administration. Linearity in pharmacokinetics was demonstrated over a 120 fold dose range. Following iv administration of resiquimod, IFNα concentrations were observed from 0.75 hpost dose, Cmax occurred at 1-2 h and the last measurable concentration was observed at approximately 3 h. A bell shaped dose response curve was observed with the maximum response observed at 0.09 mg/kg. A dose of 0.4 mg/kg achieved reproducible response results across four study days and was recommended as the challenge dose for future studies. Power analysis demonstrated that the model could be used to investigate multiple compounds at a single dose and the dose response of a single compound. Population PK/PD modelling was conducted in a sequential manner. The pharmacokinetics of resiquimod were described with a 1 compartment iv bolus model with IIV on clearance. The final PK/PD model comprised an indirect response model with stimulation of input, an effect compartment and endogenous modulator function with fixed parameter estimates for Keo, the Hill co-efficient, and M50 and IIV on Keo, EC50 and Emax. The final model did not adequately predict the IFNα time profile in individual mice and highlights that increased data may be required to provide robust estimates of the induction and elimination phase of the IFNα response. The final model did characterise the dose response relationship but predicted a reduction in response at higher doses of resiquimod when IIV was included on Emax rather than the modulator function. Primate model Following sc administration, IFNα2b had an apparent blood clearance of 2.9 mL/min/kg, an apparent volume of distribution of 3.1 L/kg and a terminal half-life of 13 h. There was low inter-animal variability in the systemic exposure (CV of 25-40%) and clearance (CV of 22-37%), however the volume and half-life demonstrated greater variability (CV of >100%)Linearity in pharmacokinetics was demonstrated over a 3 fold dose range. The pharmacokinetics were broadly comparable to those previously reported in the literature. Following IFNα2b treatment induction of neopterin and the cytokines/chemokines IL15,IL1Ra, MCP1, IL6 and eotaxin was observed. There was a sub proportional increase in the concentrations of biomarkers with the increase of IFNα2b dose from 3 MIU/kg to 10MIU/kg. An induction of body temperature was not observed. Population PK/PD modelling was conducted in a sequential manner. The pharmacokineticsof IFNα2b were described with a 2 compartment model with first order absorption and IIVon clearance and IOV on volume. A covariate analysis indicated there was no relationship with either clearance or volume with body weight. The final PK/PD model for all 6biomarkers comprised an indirect response model with stimulation of input and IIV on baseline. In addition IIV on Emax was included for neopterin, IL15, IL6 and MCP1, on Kout forIL15 and EC50 for IL1Ra and eotaxin. High values for IIV were determined for IL6 and IL1Raand the parameters for MCP1 and IL6 demonstrated poor precision. The model did not give a robust prediction of EC50 for any biomarker or Emax for IL6, MCP1 and eotaxin due to the investigated doses giving a comparable response. Conclusions Mouse Model The pharmacokinetics of resiquimod were successfully determined in the mouse for the first time following both iv infusion and iv bolus administration of resiquimod. The investigation of the dose response relationship delivered comparable data to that previously reported in the literature whereby the greatest response was observed at low doses of resiquimod and further increase in dose results in an apparent reduction in the IFNα response to a plateau. A reproducible IFNα response can be achieved between mice receiving the same resiquimod treatment across multiple study days and suggests that model that may be used in drug discovery to investigate therapeutics that have an action on the IFNα pathway. Population PK/PD modelling with the incorporation of anendogenous modulator has highlighted the gaps in the data set and improved the understanding of the model that can be used to guide future PK/PD modelling efforts. Monkey Model The pharmacokinetics of IFNα2b demonstrated that at the doses investigated in this study the receptor mediated clearance had become saturated and the clearance was driven by renal elimination and catabolism. The induction of serum neopterin concentrations andcytokines/chemokines IL15, IL6, MCP1, eotaxin and IL1Ra, which have all been implicated inthe pathogenesis of SLE, was observed in the challenged primates. A less than proportional increase in biomarker concentrations was observed with the increase in dose indicating that the concentrations of IFNα2b associated with these doses may be near the top of the concentration response curve. A population PK/PD model that describes the induction of the 6 biomarkers was developed. However, the model was limited by the small number of doses investigated. Due to the translatability of the biology between primate and humans this project has delivered a PK/PD model with disease relevant endpoints that can be used to screen compounds and potentially predict efficacious clinical doses. This project has successfully designed and validated two preclinical in vivo mechanistic PK/PD challenge models based on the hypothesis that the cytokine IFNα is central to the pathogenesis in SLE and that the mechanisms behind the induced response are translatable between species. This project has demonstrated that both models have potential in drug discovery to be used as tools to select the most appropr
Novel conducive alkaline polymer membranes for fuel cells
Get PDFA series of anion exchange membranes synthesised from poly (2, 6-dimethyl-1, 4-phenylene oxide) (PPO) have been investigated for water electrolysis applications. The prepared membranes were characterised using electrochemical impedance spectroscopy, infrared spectroscopy and thermal gravimetric analysis.Initial membranes showed very low ionic conductivity and poor stabilities (> 60 oC). Improvements to these membranes were sought by investigating the chloromethylation of PPO as well as chemical cross-linking. Cross-linking of the membranes produced a small increase in stability however these membranes remained unstable at temperatures above 70 oC. Increasing the chloromethylation reaction time to 6 hours proved to be the only way of increasing both ionic conductivity and stability, however membrane stabilities at 90 oC were still poor. Eventually a membrane stable at 90 oC for 1100 hours was prepared using a quaternised PPO and polysulphone (PS) blend.Reproducibility of the chloromethylated PPO (CMPPO) required to synthesise the membranes proved difficult. Thick polymer gels were observed which could not be used to make any membranes. A membrane was successfully prepared from a "semi gel" batch of CMPPO which showed the highest observed ionic conductivity (IC) whilst being stable at 90 oC. Improvements in stability without any loss of IC were achieved using secondary polymers as well as cross-linking agents. Investigation of a commercially available water soluble, poly(diallyldimethylammonium chloride) (polyDADMAC) proved to be ineffective due to the inability to render the materials insoluble in water. Selected membranes (based on ionic conductivity and stability) were tested for water electrolysis applications, with some of the membranes showing evolution of a small amount of oxygen and/or hydrogen.A series of anion exchange membranes synthesised from poly (2, 6-dimethyl-1, 4-phenylene oxide) (PPO) have been investigated for water electrolysis applications. The prepared membranes were characterised using electrochemical impedance spectroscopy, infrared spectroscopy and thermal gravimetric analysis.Initial membranes showed very low ionic conductivity and poor stabilities (> 60 oC). Improvements to these membranes were sought by investigating the chloromethylation of PPO as well as chemical cross-linking. Cross-linking of the membranes produced a small increase in stability however these membranes remained unstable at temperatures above 70 oC. Increasing the chloromethylation reaction time to 6 hours proved to be the only way of increasing both ionic conductivity and stability, however membrane stabilities at 90 oC were still poor. Eventually a membrane stable at 90 oC for 1100 hours was prepared using a quaternised PPO and polysulphone (PS) blend.Reproducibility of the chloromethylated PPO (CMPPO) required to synthesise the membranes proved difficult. Thick polymer gels were observed which could not be used to make any membranes. A membrane was successfully prepared from a "semi gel" batch of CMPPO which showed the highest observed ionic conductivity (IC) whilst being stable at 90 oC. Improvements in stability without any loss of IC were achieved using secondary polymers as well as cross-linking agents. Investigation of a commercially available water soluble, poly(diallyldimethylammonium chloride) (polyDADMAC) proved to be ineffective due to the inability to render the materials insoluble in water. Selected membranes (based on ionic conductivity and stability) were tested for water electrolysis applications, with some of the membranes showing evolution of a small amount of oxygen and/or hydrogen
Avoiding conflicts between the WTO agreements and the Cartagena protocol on biosafety : the principle of systemic integration and the principles that lie behind it
Get PDFGenetically modified organisms (GMOs) have been heavily traded across borders since their first commercialisation in 1996, despite the fierce global debates on their benefits and risks. International trade in GMOs are regulated mainly through the WTO Agreements and the Cartagena Protocol on Biosafety (the Protocol) at the international level. The treaties are not necessarily always consistent with one another. Their relationship also serves as a specific example of the much debated potentially conflicting relationship between trade and environment, and the particular phenomenon of the fragmentation of international law that sows the seed for conflict of international norms.;Against this background, it is pertinent to ask if there really is the potential for conflicts between the treaties. Also, how do the general international rules on conflict of norms apply to the specific relationship between the treaties? In addition, if necessary, how best might conflicts between international treaties be dealt with or avoided in general?;This thesis starts by looking at the substances of the treaties and finds that there exists the real potential for conflict. It then examines the general international rules on conflict resolution techniques, tests them on the potentially conflicting relationship between the WTO Agreements and the Protocol, and finds that existing rules could not provide definitive solutions where conflicts between the treaties arise.;It is a central argument of this thesis that conflicts between the treaties should be proactively avoided rather than resolved when disputes actually arise. More generally, with the aim of achieving sustainable development and the defragmentation of international law, the principle of systemic integration is set out as a tool which is generally used by international judicial bodies for viewing international law as a whole, as well as a viable means for avoiding conflicts between international norms.;The thesis then sketches the theoretical underpinnings of the principle of systemic integration, including the principles of mutual supportiveness, good faith, cooperation, and harmonisation, and argues that the principles that lie behind systemic integration are capable of driving integration at other levels, including institutional and domestic levels.;The thesis also includes an original empirical research undertaken in the form of interviews with state and international organisation representatives, which reaffirms and provides empirical evidence for the doctrinal arguments in this thesis.Genetically modified organisms (GMOs) have been heavily traded across borders since their first commercialisation in 1996, despite the fierce global debates on their benefits and risks. International trade in GMOs are regulated mainly through the WTO Agreements and the Cartagena Protocol on Biosafety (the Protocol) at the international level. The treaties are not necessarily always consistent with one another. Their relationship also serves as a specific example of the much debated potentially conflicting relationship between trade and environment, and the particular phenomenon of the fragmentation of international law that sows the seed for conflict of international norms.;Against this background, it is pertinent to ask if there really is the potential for conflicts between the treaties. Also, how do the general international rules on conflict of norms apply to the specific relationship between the treaties? In addition, if necessary, how best might conflicts between international treaties be dealt with or avoided in general?;This thesis starts by looking at the substances of the treaties and finds that there exists the real potential for conflict. It then examines the general international rules on conflict resolution techniques, tests them on the potentially conflicting relationship between the WTO Agreements and the Protocol, and finds that existing rules could not provide definitive solutions where conflicts between the treaties arise.;It is a central argument of this thesis that conflicts between the treaties should be proactively avoided rather than resolved when disputes actually arise. More generally, with the aim of achieving sustainable development and the defragmentation of international law, the principle of systemic integration is set out as a tool which is generally used by international judicial bodies for viewing international law as a whole, as well as a viable means for avoiding conflicts between international norms.;The thesis then sketches the theoretical underpinnings of the principle of systemic integration, including the principles of mutual supportiveness, good faith, cooperation, and harmonisation, and argues that the principles that lie behind systemic integration are capable of driving integration at other levels, including institutional and domestic levels.;The thesis also includes an original empirical research undertaken in the form of interviews with state and international organisation representatives, which reaffirms and provides empirical evidence for the doctrinal arguments in this thesis
Biochemical and molecular characterisation of the proposed P2Y₁-P2Y₁₂ receptor heterodimer in tsA201 cells
No full textThe P2Y receptor family represents a wide-ranging set of G-protein-coupled receptors that respond to purinergic ligands, and are involved in many physiological processes. From signalling assays in tsA201 cells, the Kennedy lab has previously proposed the formation of a constitutive heterodimer between co-expressed human P2Y₁ and P2Y₁₂ receptors; this could represent a target for the control of pain sensing neurones. Therefore, it was aimed in this project to characterise if the receptors physically interacted using coimmunoprecipitation,and also to investigate the endogenous expression of other potentially interacting hP2Y receptors in the tsA201 cell line. In this study, tsA201 cells were determined to express mRNA for hP2Y₁,₂,₄,₆,₁₂,₁₄ receptors using reverse transcriptase PCR and confirmatory Sanger sequencing. Furthermore, the cells demonstrated Ca2⁺ responses to all of the natural hP2Y receptor agonists tested, the order of potency being ADP > ATP > UTP > UDP. When co-expressing hP2Y₁ and hP2Y₁₂ receptors oppositely tagged with a HA or fluorescent protein (FP), immunoprecipitating for the HA tag and blotting for the FP showed a visible high molecular weight protein complex of ~130 to >250 KDa for either combination of receptors. This implied that the FP tagged receptor had co-immunoprecipitated with the other HA tagged receptor as this protein was also visible when blotting for the HA tag, signifying the presence of both tagged receptors in the complex.These results were overall suggestive of functional hP2Y₁,₂,₁₂ receptor expression in the tsA201 cell line, and possibly also hP2Y₄,₆,₁₄ receptors. The high molecular weight protein complex detected could represent a constitutive hP2Y₁ and hP2Y₁₂ receptor heterodimer as proposed. Whilst it was not possible to specifically either confirm or refute the physical formation of the heterodimer with the data acquired, it was potentially supportive, and furthermore suggested other future paths of investigation.The P2Y receptor family represents a wide-ranging set of G-protein-coupled receptors that respond to purinergic ligands, and are involved in many physiological processes. From signalling assays in tsA201 cells, the Kennedy lab has previously proposed the formation of a constitutive heterodimer between co-expressed human P2Y₁ and P2Y₁₂ receptors; this could represent a target for the control of pain sensing neurones. Therefore, it was aimed in this project to characterise if the receptors physically interacted using coimmunoprecipitation,and also to investigate the endogenous expression of other potentially interacting hP2Y receptors in the tsA201 cell line. In this study, tsA201 cells were determined to express mRNA for hP2Y₁,₂,₄,₆,₁₂,₁₄ receptors using reverse transcriptase PCR and confirmatory Sanger sequencing. Furthermore, the cells demonstrated Ca2⁺ responses to all of the natural hP2Y receptor agonists tested, the order of potency being ADP > ATP > UTP > UDP. When co-expressing hP2Y₁ and hP2Y₁₂ receptors oppositely tagged with a HA or fluorescent protein (FP), immunoprecipitating for the HA tag and blotting for the FP showed a visible high molecular weight protein complex of ~130 to >250 KDa for either combination of receptors. This implied that the FP tagged receptor had co-immunoprecipitated with the other HA tagged receptor as this protein was also visible when blotting for the HA tag, signifying the presence of both tagged receptors in the complex.These results were overall suggestive of functional hP2Y₁,₂,₁₂ receptor expression in the tsA201 cell line, and possibly also hP2Y₄,₆,₁₄ receptors. The high molecular weight protein complex detected could represent a constitutive hP2Y₁ and hP2Y₁₂ receptor heterodimer as proposed. Whilst it was not possible to specifically either confirm or refute the physical formation of the heterodimer with the data acquired, it was potentially supportive, and furthermore suggested other future paths of investigation
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